Preliminary studies indicate that lactose secretion by mammary gland slices is inhibited differently by colchicine, which blocks release into the media, than by cytochalasin B, which primarily inhibits synthesis. This suggests involvement of both microtubules and microfilaments. Based on the currently accepted paradigm for milk production by alveolar cells, inhibition could occur during synthesis of a-lactalbumin in the RER, transport of this protein to the Golgi complex, glucose uptake by the cell, lactose synthetase activity in the Golgi, transport of lactose-containing vesicles to the cell apex, and secretion by reverse pinocytosis. We intend to determine: 1) the identity and intracellular location of actin-like microfilaments in mammary gland alveolar cells using heavy meromyosin (HMM) decoration and electron microscopy of whole cells and actin extracts; 2) if tubulin is present in alveolar cells, the amount present, and the organelles with which it is associated, employing tubulin purification, gel electrophoresis, cell fractionation, and 3H-colchicine binding assays; 3) whether the changes we observed previously in mammary gland biopsies of tubulin content and assembly states in relation to pregnancy and lactation reflect changes in the alveolar cell itself, using 3H-colchicine binding assays on isolated alveolar cells; 4) if treatment with microtubule and microfilament-altering drugs produces ultastructural evidence of a block at any of the above sites; and 5) whether the floating collagen gel technique (Emerman and Pitelka, 1977) can be used to study lactogenesis in guinea pig alveolar cells and, if so, whether the synthesis and polymerization of microfilaments and microtubules regulated is by lactogenic hormones and/or cyclic nucleotides. These studies should further elucidate the respective functions of these two filamentous proteins, their interaction as a system, and their physiological regulation in milk product on.